Secondary Level Screening of Chlamydomonas Reinhardtii Mutants Defective in Circadian Gene Expression

To elucidate the signal transduction chain mediating circadian clock control, this work focuses on the isolation of Chlamydomonas reinhardtii mutants which are defective in circadian gene expression. In a previous study, the reporter gene ARS2 encoding the arylsulfatase enzyme was fused to the promoter of the circadian-regulated CABII-1 gene and transformed into the Chlamydomonas nucleus. The ble marker was introduced into the genome of this transformant via insertional mutagenesis to generate mutants defective in circadian CABII-1 expression. Potential mutants were selected based on aberrant single-point accumulative arylsulfatase activity. In this study, the arylsulfatase activity over the entire growth cycle was further investigated in these mutants and the reliability of the single-point screen was assessed. Of the 16 strains whose accumulative arylsulfatase activity did not differ from the nonmutagenized control in the single-point screen, 12 still showed no significant difference in a multiple-point screen. Of the 9 potential mutants with significant difference to the control in the single-point screen, 3 showed no significant difference in the multiple-point screen. Subsequently, 8 of the candidate mutants with aberrant reporter enzyme activity in the multiple-point screen were characterized by the abundance of their mRNA. The peak-to-trough ratio of CABII-1 and ARS2 transcript abundance was significantly reduced in 4 of these mutants.

Name thesis:degree_name

Master of Science

Discipline thesis:degree_discipline

Department of Biology

Year

2001

Author dc:creator

• Huang, Mingya

• Medical Sciences

Repository record dc:identifier

https://digitalcommons.wku.edu/theses/667

OAI identifier oai:identifier

oai:digitalcommons.wku.edu:theses-1670